Recent advances in genomics and proteomics research made numerous nucleotide and peptide sequences available, necessitating high-throughput screening of samples for presence and/or quantity of genes and/or gene expression. While automation of individual steps (e.g., DNA isolation, protein fractionation, etc.) in high-throughput screening may be performed using relatively simple instrument configurations, integration of multiple and distinct steps in automated high-throughput screening remains problematic.
For example, sample analysis for detection and quantification of one or more analytes may be performed in nano-volumes on a single chip (see e.g., “Lab-on-a-chip” from Agilent or Caliper Technologies). Such multiple analyte detection can advantageously be performed in relatively short time using minimal amounts of sample. Moreover, all steps from handling of the sample after application of the sample to detection and analysis are performed within the same device. However, identification and quantification of the detected analyte using nanoelectrophoresis is typically restricted to the size of the analyte. Moreover, resolution of individual analytes becomes increasingly difficult as the size or charge difference between the analytes decreases. Consequently, such nanoelectrophoretic systems are generally limited to characterization of an analyte by its molecular weight.
Where high resolution of molecular weights of an analyte is particularly important, analysis of complex samples may be coupled with laser desorption—time of flight mass spectroscopic analysis (see e.g., Ciphergen Biosystems' LD-TOF multi-analyte desorption chips, or Sequenome chip). Here, components of a complex sample are immobilized on a carrier chip (e.g., chip with anion exchange resin or hydrophobic interaction resin) and subjected to size analysis after desorption according to their molecular properties in an analysis system. LD-TOF coupled analysis is typically highly sensitive and often requires only minimal sample preparation. Moreover, LD-TOF coupled analysis provides relatively high resolution among particular analytes. However, identification of particular analytes is still mostly limited to size determination.
Alternatively, and especially where the analyte is a DNA or RNA, various formats of automated modular PCR-based analysis are known in the art. For example, where a single sample is analyzed for presence or absence of a particular sequence, all or almost all of the reagents and sample may be introduced into an automated system from a single cartridge (see e.g., Cepheid's i-CORE system). On the other band, and especially where a relatively high number of samples are concurrently analyzed, a full robotic PCR station may be employed (see e.g., Orchid Biocomputer SNP Analysis system). Such systems typically provide an analysis procedure that integrates sample manipulation with nucleic acid amplification and product analysis. However, automated modular PCR-based analysis typically rely on amplification of target DNA to generate appreciable signals, thereby introducing significant complexity and numerous error-prone procedures. Moreover, while PCR based systems are frequently operated in a dedicated environment using dedicated equipment to prevent non-sample specific signals, problems associated with contamination via sample carry-over may still persist. Thus, automated modular PCR-based analysis tends to be highly expensive, and is generally limited to exclusive analysis of nucleic acids.
In still further examples, nucleic acid-containing samples can be analyzed by their hybridization characteristics with at least partially complementary and immobilized nucleic acids, thereby providing quantitative and qualitative information on a particular sample. Hybridization of a nucleic acid to corresponding solid-phase immobilized nucleic acids may be controlled by variation of temperature and/or ionic strength of the environment of the nucleic acid hybrid, and there are numerous systems known in the art.
For example, high density arrays of immobilized oligonucleotides on a silicon chip may be exposed to a sample containing nucleic acids that are complementary to at least some of the immobilized oligonucleotides (see e.g., Affymetrix' GeneChip system). In such systems, a processed sample (typically a labeled and biotinylated in-vitro transcript of a previously prepared cDNA) is provided to the chip in a fluidics station that further controls flow of reagents and hybridization temperature. After complementary labeled nucleic acids have hybridized to the corresponding nucleic acids on the chip, the chip is removed from the fluidics station and manually transferred to a scanner station in which the sample is analyzed via detection of the fluorescent labels. While such analytic devices typically allow a user to determine identity, presence, and/or quantity of a vast number of DNA/RNA analytes in a sample, substantial sample preparation (typically several hours to more than one day) and hybridization times (e.g., about 16 hours at 40° C.) are frequently necessary. Moreover, analytes detected and quantified using such systems tend to be limited to nucleic acids.
Alternatively, sample capture and hybridization may be controlled via electrostatic forces (see e.g., Nanogen's NanoChip system). In such systems, capture probes and hybridization conditions may be individually controlled, thereby allowing custom addressing of individual analyte pixels. However, due to the complexity of loading and reading procedures, the analytic process is split among at least two independent devices: Analytes are typically bound in a loader section, while a reader (i.e., array processor and scanner) will perform the readout of the sample.
In another system, detection may be performed using an electronic chip that provides a signal upon binding of a signaling oligonucleotide to an analyte oligonucleotide that is bound to a corresponding oligonucleotide that is immobilized on the chip (see e.g., Motorola's iSensor system). While electronic detection and quantification may provide at least some advantages, most of such systems are prone to non-specific false-positive and/or false-negative signals due to contamination. Moreover, analytes detected and quantified using such systems tend to be limited to nucleic acids.
Thus, although various systems for micro array systems are known in the art, numerous problems still remain. Among other things, while various systems may provide at least some automation, fluid handling and sample detection/quantification of analyte binding are typically operated in separate devices, thus requiring at least some user intervention after the sample is applied to the system. Furthermore, all or almost all of the known micro array systems are limited to analysis of either nucleic acids or peptides. Therefore, there is still a need for an improved methods and systems for automated analytic devices.